The Separation And Purification Of Camellia Pericarp Polyphenols And Its Study Of Biological Activity

Camellia oleifera is a unique woody plants oil tree in southern China, wide planting area and abroad distribution. The fruit of Camellia oleifera Abel is composed of Camellia seed and Camellia shell which accounted for40%and60%of total quality, respectively. The vegetable oil squeezed from camellia seed is camellia oil that has high nutritional value and extensive biological activities, has been widely used in the fields of medicine, cosmetics etc. Although camellia shell is often used to produce tannin extract, furfural, xylitol, activated carbon, potassium carbonate and potassium pyrophosphate and other chemical products in industry and the study of its active composition is relatively rare. The separation and purification of camellia pericarp polyphenols and its effect on cell activity would be studied in this paper. The main contents and results are as follows:Camellia pericarp polyphenol(CPP)was purified by an D101resin column and SephadexLH-20gel chromatography and its purity around79.04±4.62%.The study found that CPP had strong scavenging capacity of DPPH (IC50=15.03μg/mL), ABTS·+(IC50=92.48μg/mL), O2-·(IC50=0.48mg/mL),·OH (IC50=0.66mg/mL) and inhibiting yelk peroxidation ability(IC50=0.46mg/mL). The study indicated that CPP have strong scavenging capacity of ROS radical and with the increasing concentration the removal effect was enhanced.By using agarose gel electrophoresis, the protective effect CPP and other five kinds of polyphenol monomer on DNA strand breaks induced by APPH were compared. The order from strong to weak:ECG> Quercetin> Gallic acid>CPP>EGC> EGCG.After separation and purification with macroporous resin separation, the purity of CPPCE around34.5%. By using single cell gel electrophoresis (SCGE), the final concentration of H2O2, which could induce the DNA damage of HUVEC, was determined to be50μmol/L. At this concentration, the comet cell rate was97.33%and the comet tail length was58.73±5.1μm.In50～500mg/L concentration range, CPPCE could significantly reduce the degree of DNA damage induced by H2O2and there was significant difference compared with the positive control group (P<0.01or P<0.05),which indicated CPPCE have a protective effect on DNA damage induced by H2O2.By the analysis of AGE, APPH could induce plasmid DNA stand breaks and its final concentration was10mmol/L. The study found that CPPCE had a significant protective effect on DNA strand breaks induced by APPH.The effects of different concentrations of camellia pericarp polyphenol on the extent of oxidative injury to the cells were evaluated. The viability of cells was determined by MTT assays; the level of cell DNA oxidative damage was assayed by single cell gel electrophoresis (SCGE); the apoptosis ratio of cells was detected by flow cytometry; and the expressions of p53mRNA were observed by RT-PCR; and the level of oxidative injury to HUVEC was also investigated by the measurement of malondialdehyde(MDA) levels, superoxide dismutase(SOD) levels and lactic dehydrogenase(LDH). Conclusion:camellia pericarp polyphenol can decrease the oxidative injury of endothelial cells induced by H2O2, can weaken the extent of cells DNA oxidative damage, can attenuate the expression of p53and can diminish the apoptosis of injured cells. These findings probably explain in part camellia pericarp polyphenol can prevent atherosclerotic cardiovascular diseases.